MicroRNA Let-7c-5p Protects Against Cerebral Ischemia Injury Via Mechanisms Involving The Inhibition Of Microglia Activation

Objective: We observed that Let-7c-5p, a highly conserved miRNA, was downregulated in the plasma of patients with ischemic stroke. Here, we investigated the effects of let-7c-5p on ischemic brain damage and potential mechanisms.Methods: Mouse cerebral ischemia was induced by the middle cerebral artery occlusion (MCAO). Primary microglia and BV2 microglial cells were activated in vitro by LPS, oxygen-glucose deprivation (OGD) or OGD-conditioned neuronal buffer. Acute cerebral infarction was determined by 2,3,5-triphenyltertrazolium chloride staining. Immunostaining was performed to characterize the activation of microglia. The expression of inflammatory mediators in the ischemic brain and activated microglia was analyzed with qPCR and western blotting. Luciferase reporter assay was performed to identify the site in the 3’un-translated region of the target mRNA that was targeted by let-7c-5p.Results:let-7c-5p levels were significantly decreased in the plasma of patients with ischemic stroke, as well as in mouse plasma, ipsilateral cortex and striatum at 6 and 24 hours following MCAO. Overexpression of let-7c-5p by ICV injection of miRNA mimics decreased infarction and microglia activation, as evidenced by the fact that let-7c-5p overexpression attenuated expression of pro-inflammatory mediators and ramified Iba1 immunoactivity in the ischemic brain. In vitro study further revealed that let-7c-5p was reduced in the activated microglia and let-7c-5p overexpression remarkably inhibited activation-induced expression of several pro-inflammatory mediators in microglia. Luciferase reporter assay suggest that let-7c-5p directly target the 3’-untranslated region of caspase 3 mRNA to reduce the mRNA levels of caspase 3 in microglia.Conclusions: Let-7c-5p overexpression protects against ischemic stroke by suppressing microglia possibly via downregulation of caspase 3.